The prior art discloses that deoxyribonucleic acid polymers are synthesized by attaching to a nucleotide which is immobilized via its 3′ end to a solid support in each step a nucleotide to the present or newly produced 5′ end, until an oligo- or polynucleotide of the desired length is formed. This oligo- or poly-nucleotide is then cleaved off the solid support. The solid support here is typically in a chromatographic column. The larger the column packing used here, the more difficult the process is to manage. More specifically, homogeneous packing of the column is difficult to achieve, and a powerful pump is required due to the loss of pressure along the flow path of large columns. Another problem is the edge effect which is always present in columns and which results in a chromatographic front migrating more slowly close to the wall of the column than in the center of the column, reducing the efficiency of the process.
Bonora, G. M. et al., Nucleic Acids Research, 1993, Volume 21, No. 5, pages 1213 to 1217 disclose, by way of an alternative method of synthesizing large amounts of DNA, a method for fluid-phase synthesis of oligo-nucleotides, which comprises using polyethylene glycol (PEG) as soluble support and phosphoramidite derivatives as synthons. This involves polyethylene glycol being bound to the 3′ end of a nucleoside to be extended in the synthesis reaction. After a dimethoxytrityl protective group on the nucleoside has been removed, a condensation with a nucleoside phosphoramidite occurs at the 5′ end of said nucleoside. The reaction product is precipitated and recrystallized. After capping and subsequent oxidation, the PEG-phosphate derivative obtained is precipitated, filtered and recrystallized. Said process steps of detritylation, condensation, precipitation, recrystallization, capping, oxidation, precipitation, filtration and recrystallization are then repeated until the polymer has the desired length. Finally, the polyethylene glycol is cleaved off and the oligo-nucleotide obtained is precipitated. The oligo-nucleotide is then dissolved and purified by ion exchange chromatography.
Both of the methods mentioned have the disadvantage of an exponential increase in the probability of producing an incorrect polynucleotide as a function of increasing chain length. This is due to the fact that in any one coupling or condensation step a small percentage of the nucleotide chains to be extended fails to react. Assuming a coupling efficiency of 98%, this results already in 18.3% incorrect oligomers for an oligomer formed from only 10 nucleotides. For a polymer formed from 100 nucleotides, already 86.7% of the polymers formed are incorrect. The incorrect polymers can thus greatly exceed the amount of correctly synthesized polymers. As a result, a large amount of effort is required to remove the incorrect polymers, for example by means of chromatography, in order to obtain the desired polymer in a pure form. Due to the purification required and the large amount of reagents inevitably used for producing incorrect polymers, producing a desired polymer is relatively expensive.
It is an object of the present invention to provide a method for fluid-phase synthesis of a polymer formed from monomers, by means of which method said polymer can be produced inexpensively in large quantities and with high purity.